Plant Pathogen Detection Protocols - Conventional PCR
Primers for Pectobacterium
Kang, H. W., Kwon, S. W., & Go, S. J. 2003. PCR-based specific and sensitive detection of Pectobacteriumcarotovora subsp. carotovora by primers generated from a URP-PCR fingerprinting-derived polymorphic band. Plant Pathol. 52, 127-133.
EXPCCR 5'-GCCGTAATTGCCTACCTGCTTAAG-3’
EXPCCF 5'-GAACTTCGCACCGCCGACCTTCTA-3'
INPCCR 5'-GGCCAAGCAGTGCCTGTATATCC-3'
INPCCF 5'-TTCGATC ACGCAACCTGCATTACT-3'
These primers were developed from a fragment of Pectobacterium sequence amplified by universal rice primers (URPs). The primers EXPCCR and EXPCCF amplify a 550-bp fragment from P. carotovorum and P. wasabiae strains. Digestion of this fragment with RsaI resulted in 5 different RFLPs for the P. carotovorum strains and 1 RFLP for the P. wasabiae strains. The IN primers are for use in nested PCR and amplify a 400-bp fragment.
Darrasse, A., Priou, S., Kotoujansky, A., & Bertheau, Y. 1994. PCR and restriction fragment length polymorphism of a pel gene as a tool to identify Erwinia carotovora in relation to potato diseases. Appl. Environ. Microbiol. 60, 1437-1443.
Y 1 5’-TTACCGGACG CCGAGCTGTGGCGT-3’
Y 2 5’-CAGGAAGATGTCGTTATCGCGAGT-3’
These primers amplify a fragment of the pelY family pectate lyases from Pectobacterium species, with the exception of P. betavasculorum.
De Boer, S. H. & Ward, L .J. 1995. PCR detection of Erwinia carotovora subsp. atroseptica associated with potato tissue. Phytopathol. 85, 854-858.
Eca1f 5'-CGGCATC ATAAAAACACG-3
Eca2R 5'-GCACACTTCATCCAGCGA-3'
This primer set amplifies a fragment from P. atrosepticum.
Primers for Dickeya
Nassar, A., Darrasse, A., Lemattre, M., Kotoujansky, A., Dervin, C., Vedel, R., & Bertheau, Y. (1996a) Characterization of Erwinia chrysanthemi by pectinolytic isozyme polymorphism and restriction fragment length polymorphism analysis of PCR-amplified fragments of pel genes. Appl. Environ. Microbiol. 62, 2228-2235.
ADE1 5’-GATCAGAAAGCCCGCAG CCAGAT-3’
ADE2 5’-CTGTGGCCGATCAGGATGGTTTTGTCGTGC-3'
This primer set amplifies a 420-bp pectate lyase gene fragment from all Dickeya strains tested, but not from any other soft rot enterobacterial species .